UMR3244 – Dynamique de l’information génétique

Publications de l’équipe

Année de publication : 2007

Valérie Borde (2007 Aug 4)

The multiple roles of the Mre11 complex for meiotic recombination.

Chromosome research : an international journal on the molecular, supramolecular and evolutionary aspects of chromosome biology : 551-63 En savoir plus
Résumé

During the first meiotic prophase, numerous DNA double-strand breaks (DSB) are formed in the genome in order to initiate recombination between homologous chromosomes. The conserved Mre11 complex, formed of Mre11, Rad50 and Nbs1 (Xrs2 in Saccharomyces cerevisiae) proteins, plays a crucial role in mitotic cells for sensing and repairing DSB. In meiosis the Mre11 complex is also required for meiotic recombination. Depending on the organisms, the Mre11 complex is required for the formation of the DSB catalysed by the transesterase Spo11 protein. It then plays a unique function in removing covalently attached Spo11 from the 5′ extremity of the breaks through its nuclease activity, to allow further break resection. Finally, the Mre11 complex also plays a role during meiosis in bridging DNA molecules together and in sensing Spo11 DSB and activating the DNA damage checkpoint. In this article the different biochemical functions of the Mre11 complex required during meiosis are reviewed, as well as the consequences of Mre11 complex inactivation for meiosis in several organisms. Finally, I describe the meiotic phenotypes of several animal models that have been developed to model hypomorphic mutations of the Mre11 complex, involved in humans in some genetic instability disorders.

Replier
Franck Toledo, Olivier Bluteau, Iva Simeonova (2007 Jul 17)

[The activation of p53 in tumors: a promising strategy against cancer].

Médecine sciences : M/S : 565-7 En savoir plus
Résumé

Replier
Franck Toledo, Geoffrey M Wahl (2007 May 15)

MDM2 and MDM4: p53 regulators as targets in anticancer therapy.

The international journal of biochemistry & cell biology : 1476-82 En savoir plus
Résumé

The gene TP53, encoding transcription factor p53, is mutated or deleted in half of human cancers, demonstrating the crucial role of p53 in tumor suppression. Importantly, p53 inactivation in cancers can also result from the amplification/overexpression of its specific inhibitors MDM2 and MDM4 (also known as MDMX). The presence of wild-type p53 in those tumors with MDM2 or MDM4 overexpression stimulates the search for new therapeutic agents to selectively reactivate it. This short survey highlights recent insights into MDM2 and MDM4 regulatory functions and their implications for the design of future p53-based anticancer strategies. We now know that MDM2 and MDM4 inhibit p53 in distinct and complementary ways: MDM4 regulates p53 activity, while MDM2 mainly regulates p53 stability. Upon DNA damage, MDM2-dependent degradation of itself and MDM4 contribute significantly to p53 stabilization and activation. These and other data imply that the combined use of MDM2 and MDM4 antagonists in cancer cells expressing wild-type p53 should activate p53 more significantly than agents that only antagonize MDM2, resulting in more effective anti-tumor activity.

Replier

Année de publication : 2006

Nicolas Robine, Norio Uematsu, Franck Amiot, Xavier Gidrol, Emmanuel Barillot, Alain Nicolas, Valérie Borde (2006 Dec 26)

Genome-wide redistribution of meiotic double-strand breaks in Saccharomyces cerevisiae.

Molecular and cellular biology : 1868-80 En savoir plus
Résumé

Meiotic recombination is initiated by the formation of programmed DNA double-strand breaks (DSBs) catalyzed by the Spo11 protein. DSBs are not randomly distributed along chromosomes. To better understand factors that control the distribution of DSBs in budding yeast, we have examined the genome-wide binding and cleavage properties of the Gal4 DNA binding domain (Gal4BD)-Spo11 fusion protein. We found that Gal4BD-Spo11 cleaves only a subset of its binding sites, indicating that the association of Spo11 with chromatin is not sufficient for DSB formation. In centromere-associated regions, the centromere itself prevents DSB cleavage by tethered Gal4BD-Spo11 since its displacement restores targeted DSB formation. In addition, we observed that new DSBs introduced by Gal4BD-Spo11 inhibit surrounding DSB formation over long distances (up to 60 kb), keeping constant the number of DSBs per chromosomal region. Together, these results demonstrate that the targeting of Spo11 to new chromosomal locations leads to both local stimulation and genome-wide redistribution of recombination initiation and that some chromosomal regions are inherently cold regardless of the presence of Spo11.

Replier
Franck Toledo, Crystal J Lee, Kurt A Krummel, Luo-Wei Rodewald, Chung-Wen Liu, Geoffrey M Wahl (2006 Dec 13)

Mouse mutants reveal that putative protein interaction sites in the p53 proline-rich domain are dispensable for tumor suppression.

Molecular and cellular biology : 1425-32 En savoir plus
Résumé

The stability and activity of tumor suppressor p53 are tightly regulated and partially depend on the p53 proline-rich domain (PRD). We recently analyzed mice expressing p53 with a deletion of the PRD (p53(DeltaP)). p53(DeltaP), a weak transactivator hypersensitive to Mdm2-mediated degradation, is unable to suppress oncogene-induced tumors. This phenotype could result from the loss of two motifs: Pin1 sites proposed to influence p53 stabilization and PXXP motifs proposed to mediate protein interactions. We investigated the importance of these motifs by generating mice encoding point mutations in the PRD. p53(TTAA) contains mutations suppressing all putative Pin1 sites in the PRD, while p53(AXXA) lacks PXXP motifs but retains one intact Pin1 site. Both mutant proteins accumulated in response to DNA damage, although the accumulation of p53(TTAA) was partially impaired. Importantly, p53(TTAA) and p53(AXXA) are efficient transactivators and potent suppressors of oncogene-induced tumors. Thus, Pin1 sites in the PRD may modulate p53 stability but do not significantly affect function. In addition, PXXP motifs are not essential, but structure dictated by the presence of prolines, PXXXXP motifs that may mediate protein interactions, and/or the length of this region appears to be functionally significant. These results may explain why the sequence of the p53 PRD is so variable in evolution.

Replier
Franck Toledo, Geoffrey M Wahl (2006 Nov 28)

Regulating the p53 pathway: in vitro hypotheses, in vivo veritas.

Nature reviews. Cancer : 909-23 En savoir plus
Résumé

Mutations in TP53, the gene that encodes the tumour suppressor p53, are found in 50% of human cancers, and increased levels of its negative regulators MDM2 and MDM4 (also known as MDMX) downregulate p53 function in many of the rest. Understanding p53 regulation remains a crucial goal to design broadly applicable anticancer strategies based on this pathway. This Review of in vitro studies, human tumour data and recent mouse models shows that p53 post-translational modifications have modulatory roles, and MDM2 and MDM4 have more profound roles for regulating p53. Importantly, MDM4 emerges as an independent target for drug development, as its inactivation is crucial for full p53 activation.

Replier
Maria Antonietta Cerone, J Arturo Londoño-Vallejo, Chantal Autexier (2006 Aug 8)

Telomerase inhibition enhances the response to anticancer drug treatment in human breast cancer cells.

Molecular cancer therapeutics : 1669-75 En savoir plus
Résumé

Breast cancer is the most common malignancy among women. Current therapies for breast tumors are based on the use of chemotherapeutic drugs that are quite toxic for the patients and often result in resistance. Telomerase is up-regulated in 95% of breast carcinomas but not in adjacent normal tissues. Therefore, it represents a very promising target for anticancer therapies. Unfortunately, the antiproliferative effects of telomerase inhibition require extensive telomere shortening before they are fully present. Combining telomerase inhibition with common chemotherapeutic drugs can be used to reduce this lag phase and induce tumor cell death more effectively. Few studies have analyzed the effects of telomerase inhibition in combination with anticancer drugs in breast cancer cells. In this study, we inhibited telomerase activity in two breast cancer cell lines using a dominant-negative human telomerase reverse transcriptase and analyzed cell viability after treatment with different anticancer compounds. We found that dominant-negative human telomerase reverse transcriptase efficiently inhibits telomerase activity and causes telomere shortening over time. Moreover, cells in which telomerase was suppressed were more sensitive to anticancer agents independently of their mechanism of action and this sensitization was dependent on the presence of shorter telomeres. Altogether, our data show that blocking telomere length maintenance in combination with anticancer drugs can be used as an effective way to induce death of breast cancer cells.

Replier
J Graakjaer, J A Londono-Vallejo, K Christensen, S Kølvraa (2006 Jun 29)

The pattern of chromosome-specific variations in telomere length in humans shows signs of heritability and is maintained through life.

Annals of the New York Academy of Sciences : 311-6 En savoir plus
Résumé

This paper characterizes the distribution of telomere length on individual chromosome arms in humans. By fluorescent in situ hybridization (FISH), followed by computer-assisted analysis of digital images, it is shown that the distribution of telomere length on individual chromosome arms is not random, but that humans have a common telomere profile. This profile exists in lymphocytes, amniocytes and fibroblasts, and seems to be conserved during life. A closer look at the overall pattern of the profile shows that the length of the telomeres in general follows the total chromosome length. In addition to the common profile, it is found that each person has specific characteristics, which are also conserved throughout life. Studying both twins and families we have obtained indications that these individual characteristics are at least partly inherited. Altogether, our results suggest that the length of individual telomeres might occasionally play a role in the heritability of life span.

Replier
M A Cerone, J A Londoño-Vallejo, C Autexier (2006 Jun 13)

Mutated telomeres sensitize tumor cells to anticancer drugs independently of telomere shortening and mechanisms of telomere maintenance.

Oncogene : 7411-20 En savoir plus
Résumé

Telomerase is a ribonucleoprotein complex that maintains the stability of chromosome ends and regulates replicative potential. Telomerase is upregulated in over 85% of human tumors, but not in adjacent normal tissues and represents a promising target for anticancer therapy. Most telomerase-based therapies rely on the inhibition of telomerase activity and require extensive telomere shortening before inducing any antiproliferative effect. Disturbances of telomere structure rather than length may be more effective in inducing cell death. Telomerase RNA subunits (hTRs) with mutations in the template region reconstitute active holoenzymes that incorporate mutated telomeric sequences. Here, we analysed the feasibility of an anticancer approach based on the combination of telomere destabilization and conventional chemotherapeutic drugs. We show that a mutant template hTR dictates the synthesis of mutated telomeric repeats in telomerase-positive cancer cells, without significantly affecting their viability and proliferative ability. Nevertheless, the mutant hTR increased sensitivity to anticancer drugs in cells with different initial telomere lengths and mechanisms of telomere maintenance and without requiring overall telomere shortening. This report is the first to show that interfering with telomere structure maintenance in a telomerase-dependent manner may be used to increase the susceptibility of tumor cells to anticancer drugs and may lead to the development of a general therapy for the treatment of human cancers.

Replier
J-C Marine, S Francoz, M Maetens, G Wahl, F Toledo, G Lozano (2006 Mar 18)

Keeping p53 in check: essential and synergistic functions of Mdm2 and Mdm4.

Cell death and differentiation : 927-34 En savoir plus
Résumé

Replier
Jesper Graakjaer, Héra Der-Sarkissian, Annette Schmitz, Jan Bayer, Gilles Thomas, Steen Kolvraa, José-Arturo Londoño-Vallejo (2006 Jan 28)

Allele-specific relative telomere lengths are inherited.

Human genetics : 344-50 En savoir plus
Résumé

Previous studies have indicated that single relative telomere lengths are defined in the zygote. In order to explore the possibility that single telomere lengths segregate in families, we compared relative telomere lengths obtained from allelic chromosome extremities transmitted from parent to child, representing a total of 31 independent meiotic events. We find a significant positive correlation of 0.65 (P=0.0004) between these telomere lengths, whereas the correlation between the non-transmitted parental homologue and the transmitted homologue in the child is not statistically significant (r=0.16; P=0.195). This study indicates that, even though there is a telomerase-mediated maintenance/elongation of telomeres in germ cells, allele-specific relative telomere lengths are preserved in the next generation.

Replier

Année de publication : 2005

Fred Goldman, Rachida Bouarich, Shashikant Kulkarni, Sara Freeman, Hong-Yan Du, Lea Harrington, Philip J Mason, Arturo Londoño-Vallejo, Monica Bessler (2005 Nov 15)

The effect of TERC haploinsufficiency on the inheritance of telomere length.

Proceedings of the National Academy of Sciences of the United States of America : 17119-24 En savoir plus
Résumé

Telomeres protect chromosome ends from end-to-end fusion and degradation. Loss of telomere function causes cell-cycle arrest or cell death. Autosomal dominant dyskeratosis congenita (AD DC), a rare inherited bone marrow failure syndrome, is caused by mutations in TERC, the RNA component of telomerase. Here, we studied the telomere dynamics over three generations in a 32-member extended family with AD DC due to a TERC gene deletion. Our analysis shows that peripheral blood cells from family members haploinsufficient for TERC have very short telomeres. Telomeres are equally short in all individuals carrying the TERC gene deletion irrespective of their age. Chromosome-specific telomere analysis distinguishing the parental origin of telomeres showed that in gene deletion carriers, paternal and maternal telomeres are similarly short and similar in length to those of the affected parent. In children of affected parents who have normal TERC genes, parental telomeres are again similar in length, but two generations appear to be necessary to fully restore normal telomere length. These results are consistent with a model in which telomerase preferentially acts on the shortest telomeres. When TERC is limiting, this preference leads to the accelerated shortening of longer telomeres. The limited amount of active telomerase in TERC RNA haploinsufficiency may not be able to maintain the minimal length of the increasing number of short telomeres. Thus, the number of cells with excessively short telomeres and the degree of residual telomerase activity may determine the onset of disease in patients with AD DC.

Replier
Delphine T Marie-Egyptienne, Maria Antonietta Cerone, J Arturo Londoño-Vallejo, Chantal Autexier (2005 Sep 30)

A human-Tetrahymena pseudoknot chimeric telomerase RNA reconstitutes a nonprocessive enzyme in vitro that is defective in telomere elongation.

Nucleic acids research : 5446-57 En savoir plus
Résumé

The phylogenetically-derived secondary structures of telomerase RNAs (TR) from ciliates, yeasts and vertebrates are surprisingly conserved and contain a pseudoknot domain at a similar location downstream of the template. As the pseudoknot domains of Tetrahymena TR (tTR) and human TR (hTR) mediate certain similar functions, we hypothesized that they might be functionally interchangeable. We constructed a chimeric TR (htTR) by exchanging the hTR pseudoknot sequences for the tTR pseudoknot region. The chimeric RNA reconstituted human telomerase activity when coexpressed with hTERT in vitro, but exhibited defects in repeat addition processivity and levels of DNA synthesis compared to hTR. Activity was dependent on tTR sequences within the chimeric RNA. htTR interacted with hTERT in vitro and dimerized predominantly via a region of its hTR backbone, the J7b/8a loop. Introduction of htTR in telomerase-negative cells stably expressing hTERT did not reconstitute an active enzyme able to elongate telomeres. Thus, our results indicate that the chimeric RNA reconstituted a weakly active nonprocessive human telomerase enzyme in vitro that was defective in telomere elongation in vivo. This suggests that there may be species-specific requirements for pseudoknot functions.

Replier
Maria A Cerone, Chantal Autexier, J Arturo Londoño-Vallejo, Silvia Bacchetti (2005 Aug 24)

A human cell line that maintains telomeres in the absence of telomerase and of key markers of ALT.

Oncogene : 7893-901 En savoir plus
Résumé

In human somatic cells proliferation results in telomere shortening due to the end replication problem and the absence of adequate levels of telomerase activity. The progressive loss of telomeric DNA has been associated with replicative senescence. Maintenance of telomere structure and function is, therefore, an essential requisite for cells that proliferate indefinitely. Human cells that have acquired the immortal phenotype mostly rely on telomerase to compensate for telomere shortening with cell division. However, a certain percentage of immortalized cell lines and human tumors maintain their telomeres by Alternative Lengthening of Telomeres (ALT), a mechanism not fully understood but apparently based on homologous recombination. Here, we report the isolation of an immortal human cell line that is derived from an ALT cell line but maintains telomeres in the absence of key features of ALT and of telomerase. The properties of these cells suggest that the identification of ALT cells may not be reliably based on known ALT markers. This finding is of relevance for discriminating between the mortal and immortal phenotype among telomerase-negative cells in vitro and in vivo, particularly in regard to the development of pharmacological approaches for cancer treatment based on telomerase inhibition.

Replier
Maria Antonietta Cerone, Ryan J Ward, J Arturo Londoño-Vallejo, Chantal Autexier (2005 Mar 9)

Telomerase RNA mutated in autosomal dyskeratosis congenita reconstitutes a weakly active telomerase enzyme defective in telomere elongation.

Cell cycle (Georgetown, Tex.) : 585-9 En savoir plus
Résumé

Dyskeratosis congenita (DC) is a rare multi-system syndrome characterized by nail dystrophy, abnormal skin pigmentation and mucosal leukoplakia. The gene mutated in the X-linked form of human DC encodes for dyskerin, a nucleolar pseudourydilase that is involved in rRNA maturation. Dyskerin is also involved in telomerase function through its interaction with the telomerase RNA (hTR). Mutations in dyskerin result in low levels of hTR, decreased telomerase activity and telomere shortening. Autosomal dominant DC is characterized by mutations in hTR, supporting the hypothesis that the DC phenotype may be caused by impaired telomere maintenance. Several mutations have been identified in different regions of hTR in patients affected by autosomal dominant DC. Recent reports have shown that coexpression of wild-type hTR with hTR harboring mutations found in the pseudoknot domain does not affect telomerase activity in vitro. However, these studies did not assess the consequences of mutant hTR expression at the telomeres. Here we provide the first direct in vivo evidence that a mutant hTR carrying the GC to AG double substitution in the pseudoknot at nucleotides 107-108 found in patients affected by autosomal dominant DC does not behave as a dominant-negative for telomere maintenance. Rather it reconstitutes a weakly active telomerase enzyme, which is defective in telomere elongation.

Replier